Probes for INS

The peroxisome proliferator-activated receptor (PPAR) nuclear receptor has been an interesting target for the treatment of chronic diseases. Although the efficacy of PPAR pan agonists in several metabolic diseases has been well studied, the effect of PPAR pan agonists on kidney fibrosis development has not been demonstrated. To evaluate the effect of the PPAR pan agonist MHY2013, a folic acid (FA)-induced in vivo kidney fibrosis model was used. MHY2013 treatment significantly controlled decline in kidney function, tubule dilation, and FA-induced kidney damage. The extent of fibrosis determined using biochemical and histological methods showed that MHY2013 effectively blocked the development of fibrosis. Pro-inflammatory responses, including cytokine and chemokine expression, inflammatory cell infiltration, and NF-κB activation, were all reduced with MHY2013 treatment. To demonstrate the anti-fibrotic and anti-inflammatory mechanisms of MHY2013, in vitro studies were conducted using NRK49F kidney fibroblasts and NRK52E kidney epithelial cells. In the NRK49F kidney fibroblasts, MHY2013 treatment significantly reduced TGF-β-induced fibroblast activation. The gene and protein expressions of collagen I and α-smooth muscle actin were significantly reduced with MHY2013 treatment. Using PPAR transfection, we found that PPARγ played a major role in blocking fibroblast activation. In addition, MHY2013 significantly reduced LPS-induced NF-κB activation and chemokine expression mainly through PPARβ activation. Taken together, our results suggest that administration of the PPAR pan agonist effectively prevented renal fibrosis in both in vitro and in vivo models of kidney fibrosis, implicating the therapeutic potential of PPAR agonists against chronic kidney diseases.

Sprouty1 (Spry1) regulates the differentiation of vascular smooth muscle cells (VSMC), and our aim was to determine its role in atherogenesis. A significant proportion of cells within atherosclerotic lesions are derived from migration and pathological adaptation of medial VSMC.We used global Spry1 null mouse, and Myh11-CreERT2, ROSA26-STOPfl/fl-tdTomato-Spry1fl/fl mice to allow for lineage tracing and conditional Spry1 deletion in VSMC. Atherosclerosis was induced by injection of a mutant form of mPCSK9D377Y-AAV followed by Western diet. Human aortic VSMC (hVSMC) with shRNA targeting of Spry1 were also analyzed.Global loss of Spry1 increased inflammatory markers ICAM1 and Cox2 in VSMC. Conditional deletion of Spry1 in VSMC had no effect on early lesion development, despite increased Sca1high cells. After 26 weeks of Western diet, mice with VSMC deletion of Spry1 had increased plaque burden, with reduced collagen content and smooth muscle alpha actin (SMA) in the fibrous cap. Lineage tracing via tdTomato marking Cre-recombined cells indicated that VSMC with loss of Spry1 had decreased migration into the lesion, noted by decreased proportions of tdTomato+ and tdTomato+/SMA + cells. Loss-of-function of Spry1 in hVSMC increased mesenchymal and activation markers, including KLF4, PDGFRb, ICAM1, and Cox2. Loss of Spry1 enhanced the effects of PDGFBB and TNFa on hVSMC.Loss of Spry1 in VSMC aggravated plaque formation at later stages, and increased markers of instability. Our results indicate that Spry1 suppresses the mesenchymal and inflammatory phenotype of VSMC, and its expression in VSMC is protective against chronic atherosclerotic disease.

A high-fat diet (HFD) is a major risk factor for chronic kidney disease. Although HFD promotes renal injury, characterized by increased inflammation and oxidative stress leading to fibrosis, the underlying mechanism remains elusive. Here, we investigated the role and mechanism of protease-activating receptor 2 (PAR2) activation during HFD-induced renal injury in C57/BL6 mice. HFD for 16 weeks resulted in kidney injury, manifested by increased blood levels of blood urea nitrogen, increased levels of oxidative stress with inflammation, and structural changes in the kidney tubules. HFD-fed kidneys showed elevated PAR2 expression level in the tubular epithelial region. To elucidate the role of PAR2, PAR2 knockout mice and their littermates were administered HFD. PAR2 deficient kidneys showed reduced extent of renal injury. PAR2 deficient kidneys showed significantly decreased levels of inflammatory gene expression and macrophage infiltration, followed by reduced accumulation of extracellular matrix proteins. Using NRK52E kidney epithelial cells, we further elucidated the mechanism and role of PAR2 activation during renal injury. Palmitate treatment increased PAR2 expression level in NRK52E cells and scavenging of oxidative stress blocked PAR2 expression. Under palmitate-treated conditions, PAR2 agonist-induced NF-κB activation level was higher with increased chemokine expression level in the cells. These changes were attenuated by the depletion of oxidative stress. Taken together, our results suggest that HFD-induced PAR2 activation is associated with increased levels of renal oxidative stress, inflammatory response, and fibrosis.